Considerable interest exists in the application of genetic engineering techniques for the production of commercially valuable products such as insulin, human and animal growth hormones and enzymes. Much of the work to date has involved use of Escherichia coli as the host into which foreign genetic material is introduced. Expression of the genetic material in E. coli results in production of desired products. When combined with growth of genetically engineered cells in culture, it permits production of the desired products in commercially meaningful yields. Unfortunately, use of E. coli as a host is associated with certain disadvantages. As a result, alternative hosts, including other bacteria and yeast, are under investigation.
One particularly promising host for commercial applications of genetic engineering is Bacillus subtilis. B. subtilis is a non-pathogenic, gram positive bacterium which is eaten daily by millions of Japanese as part of a fermented soybean product. B. subtilis may be the safest bacterium in which to achieve expression of foreign genes whose products, e.g. interferon, will be purified and subsequently injected into humans for at least two reasons. First, B. subtilis is known to be non-pathogenic. Secondly, E. coli is known to produce endotoxins which may contaminate genetic products and induce endotoxic shock in humans.
Direct expression in Bacillus subtilis of a gene originating in Escherichia coli has been achieved in a single reported case [Rubin, E. M., et al., Gene 10:227-235 (1980)] There the E. coli gene specifying thymidylate synthetase expressed upon integration into the B. subtilis chromosome. However, E. coli genes residing on plasmids typically do not express at the level of genetic function in B. subtilis. [Kreft, J., et al., Mol. Gen. Genet. 162:59-67 (1978)]. The least complex explanation for the lack of foreign gene expression in B. subtilis is an absence of correct transcription and/or translation. In vitro transcription studies have indicated that E. coli RNA polymerase is significantly more efficient in initiating transcription from an E. coli promoter than is the B. subtilis RNA polymerase [Lee, G., et al., Mol Gen. Genet. 180:57-65 (1980)]. It was therefore suspected that inserting an E. coli gene, or other foreign gene, into a segment of DNA known to be efficiently transcribed in B. subtilis might permit functional expression of the foreign DNA.